The objectives of this project are, first, to develop less toxic immunosuppressive conditioning programs for allogeneic hematopoietic stem cell transplants (HSCT) in a canine model that can be applied to patients who do not have HLA-identical sibling donors and, second, to refine existing regimens for major histocompatibility complex matched recipients. In the last grant period, we described a regimen in DLA- identical matched canine littermates consisting of a non-marrow ablative dose of 200 cGy total body irradiation (TBI) before, and mycophenolate mofetil (MMF) and cyclosporine (CSP) after HSCT, which we successfully translated to human patients with malignant and non- malignant diseases. The principal underlying the new transplant approach was to use post-grafting immunosuppression with MMF/CSP, not only for better control of graft-versus-host disease (GVHD), but also to suppress host-versus-graft reactions. This way, the intensity of the conditioning programs could be reduced. Combing the new principal with previously published observations, we have shown that DLA- haploidentical HSCT could be almost uniformly established after only 450 cGy TBI when drugs were given, in addition, an anti-CD44 monoclonal antibody (Mab, S5) before transplant. We plan to lower the TBI dose to the sublethal range of 200 cGy and to obtain better control of GVHD in this model by evaluating three distinctly different approaches: (1) extending post-grafting MMF/CSP from the previously used 28 and 35 days, respectively to 100 days; (2) evaluating an antagonist to LFA1 (leukocyte functional antigen-1) peritransplant; and (3) adding T cell co- stimulatory blockade with CTLA4lg+/- an Mab to CD154 (CD40 ligand) to the regimen. In DLA-identical littermates, we will examine whether targeted irradiation firm a short-lived alpha-emitting radionuclide, Bismuth 213 (Bi-213), delivered through appropriate MAbs can be substituted for 200 cGy external beam TBI in providing immunosuppression necessary for engraftment. We will first target host cells with a pan-hematopoietic anti-CD45 antibody, and then test T cell- specific ablation with an anti-T cell receptor antibody. Successful strategies will be used in combination with the goal of completely replacing TBI with less toxic immunosuppression regimens.